My laboratory has been focusing on gene expression and posttranscriptional regulation of oncogenic DNA tumor viruses, using papillomaviruses and Kaposi's sarcoma-associated herpesvirus (KSHV) as our models to understand how a tumor virus expresses its gene for virus replication and for oncogenesis. Unlike most mammalian genes, both papillomavirus and KSHV genomes feature much more unique split genes and commonly encodes their genes, including several viral oncogenes, with significant numbers of the bicistronic or polycistronic transcripts that undergo extensive alternative splicing. Because viral oncogenesis is one of the top priorities of the HIV and AIDS Malignancy Branch of CCR, NCI, the goal of my research program is to understand the molecular details regulating expression of oncogenic and/or pathogenic viral genes and their regulation at posttranscriptional levels.Human papillomavirus type 16 (HPV16) infection, acquired via sexual transmission, is widely recognized as a leading cause of cervical cancer. High prevalence and incidence of cervical HPV infection has been observed among HIV+ women and cervical cancer has been the most common malignancy among women with AIDS in both Europe and the USA. Two viral oncoproteins, E6 and E7, of HPV16 are involved in cervical carcinogenesis and respectively, inactivate cellular tumor suppressor proteins p53 and pRb. HPV16 E6 and E7 are transcribed as a single bicistronic RNA bearing 3 exons and 2 introns, with the intron 1 in the E6 coding region. It has been believed that the E6 is expressed from the bicistronic RNA, without splicing of the intron 1. This has raised several important questions on how this intron escapes splicing, what regulates this escape, and how an RNA containing an intron is exported to cytoplasm to translate E6 protein. We have studied the mechanisms involved in this splicing regulation and found that the intron 1 splicing subjects to exon definition (size) by a cap structure on the RNA 5' end. Escaping splicing of the intron 1 results in production of unspliced E6 mRNA to encode viral oncoprotein E6, whereas splicing of the intron promotes E7 oncoprotein expression. This has been evidenced by multiple approaches in transient transfection and in cervical cancer-derived HPV16+ and HPV18+ cell lines. Recently, we have further demonstrated that some high-risk HPV proteins, such as HPV16 E2 and E6, are RNA binding proteins in vitro that interact with cellular splicing factors and suppress viral RNA splicing of HPV16 E6E7 pre-mRNAs. This observation appears to help us understand how a high-risk HPV control E6 oncogene expression. Currently, we are focusing on (1) expression of HPV16 E2 and E6 in vivo to examine their function in RNA binding and splicing regulation in mammalian cells; (2) identification of a novel spliced isoform of E6 and its biological function; (3) control of HPV16 E6 and E7 expression by siRNA targeting to the bicistronic E6E7 RNA transcripts; (4) microRNA expression and regulation in cervical cancer. These studies will lead us to better understand oncogenesis of HPV16 in development of cervical cancer.KSHV infection has been associated with all forms of Kaposi's sarcoma (KS), primary effusion lymphoma [PEL], and multicentric Castleman's disease and displays two viral life cycles. Among those malignancies, KS occurs frequently in patients infected with HIV. Latent KSHV infection in KS lesions and PEL-derived B cells features the highly restricted expression of only five viral genes. The lytic KSHV infection can be induced by chemicals in PEL-derived B cells with latent KSHV infection. In this lytic switch, a KSHV transactivator, ORF50, is absolutely required. ORF50 is an immediate-early gene transcribed as a polycistronic RNA with 5 exons and 4 introns.